It is becoming evident that hemorrhagic shock (HS) initiates inflammatory cascades and that these cascades participate in organ damage. Unlike sepsis, the systemic inflammation after HS is not due to a specific site of infection. Instead, processes initiated by reduced perfusion and reperfusion are responsible. Bacteria or bacterial products (i.e., lipopolysaccharide, LPS) released from the injured gut are suspected inflammatory stimuli in HS. LPS has been shown to activate responsive cells through the interaction of LPS with either soluble or membrane-bound CD14 in a process that is accelerated by LPS-binding protein. We have found that CD14 expression is markedly upregulated during shock with further upregulation following resuscitation. This includes a dramatic increase in CD14 on cells not previously shown to express CD14 (e.g., respiratory epithelium). We propose to address three key questions to define the role of LPS and LPS to CD14 interaction in inflammation and organ injury following HS. These questions include: 1) How is CD14 upregulated in shock? 2) Does LPS to CD14 interaction promote inflammation or injury in HS? and 3) Does the increase in lung CD14 expression result in an LPS hypersensitive state following HS? We will take advantage of our expertise in rodent HS and ischemia/reperfusion models to utilize several unique resources to fully address these questions. Key resources include CD14 and LBP knockout mice, novel LPS antagonist, animals which overexpress CD14, and previously unavailable rat-specific reagents. Under AIM I: To determine the mechanism of the upregulation of CD14 in hemorrhagic shock, we will determine the mechanisms of upregulation of CD14 in the respiratory epithelium using animals subjected to HS, subcutaneously implanted trachea, and cultured epithelial cells. AIM II: To deter-mine the functional role of CD14 in the inflammatory changes and tissue injury following resuscitation from hemorrhagic shock, will specifically focus on the role of CD14 in the initiation of inflammation. This will be accomplished using transgenic/knockout animals subjected to HS and animals pretreated with neutralizing anti-CD14 antibodies or LPS antagonists. AIM m: To determine the role of CD14 In endotoxin hypersensitivity following hemorrhagic shock, will establish whether increases in CD14 expression confer a heightened sensitivity to LPS using the same in vitro and in vivo models. These experiments should yield important data on the mechanisms of shock-induced gene expression and provide definitive answers on the role of LPS in HS-mediated inflammation and organ damage.